The unique class of broad host range plasmids (P-1 incompatibility group) and the phages dependent upon them will be studied. To understand the host range capability, these experiments seek to determine the nature of the functions required to replicate in a variety of bacterial hosts and the means by which the genes for these functions are expressed and controlled in each host. Replication regions of the Inc P-l plasmid RK2 will be cloned in phage lambda to map the genes precisely, isolate mutants, identify gene products and determine their function. These genes will be used to probe other Inc P-l plasmids for the presence of similar coding regions, and for variations in gene arrangement which may have consequences in regulation. Functional similarity will be determined by complementation studies. The temporal sequence of plasmid gene transcription will be examined with the lambda-hybrids. The effect of mutations and inhibitors will be important to understanding mechanisms of control. E. coli RNA polymerase binding sites will be mapped on the RK2 genome to determine if the natural host polymerase is adequate for expression of the plasmid genes. The genetic interaction of the broad host range phages and the plasmids upon which they depend will be examined in more detail. These phages will be used to compare gene expression in different hosts. The properties of the RK2 trans-acting replication functions will be used to develop a unique cloning vehicle system.